Adequate muscle mass and function are critical for human health. Skeletal muscle comprises ~50-60% of total body mass and is one of the major tissues involved in regulating metabolism, locomotion and strength. Loss of muscle mass results in weakness and impaired motility, and if severe enough, in increased morbidity and mortality. Therefore, defining the molecular and cellular pathways that regulate growth of skeletal muscle is critical for human health. Our lab has shown non-redundant roles for different members of the NFAT family of transcription factors in skeletal muscle growth, specifically the NFATc2 and NFATc3 isoforms. Recently a new isoform of NFAT, NFAT5, has been discovered, which displays many differences from the conventional NFAT family members. Our recent data indicates NFAT5 is expressed in muscle cells and regulates multiple phases of myogenesis. In addition, our preliminary data show that creatine, an activator of NFAT5 transcriptional activity, enhances muscle growth. Furthermore, downstream targets of NFAT5 signaling regulate myogenesis. The overall goal of this proposal is to determine how the NFAT5 signaling pathway regulates muscle growth using an integrated approach in vivo and in vitro. A set of 4 aims are proposed to analyze NFAT5 function during muscle growth during muscle regeneration in vivo (Aim 1), determine the role of NFAT5 in regulating myoblast proliferation, migration, differentiation and fusion in vitro (Aim 2), analyze the NFAT5-dependent and -independent roles of creatine on muscle growth (Aim 3) and study the role of a newly identified NFAT5 target gene in myogenesis (Aim 4). A combination of cellular and molecular approaches will be utilized. Our proposed studies will define a novel pathway in muscle for regulating muscle growth. Understanding the role of the NFAT5 pathway in regulating muscle size may lead to new strategies for manipulating muscle cells in disease, repair and aging and define novel therapeutic targets for enhancing muscle growth.